If an association between the widely used pesticide, endosulfan and male infertility has been found in the case of humans living in areas where it has been extensively used, laboratory studies on mice have now confirmed that the pesticide indeed causes male infertility.
A significant reduction in sperm count and sperm motility in mice was found to cause infertility in mice. The pesticide-induced infertility was found to be male specific. The results were recently published in the journal Cell Death and Disease.
“Thirty to 33 per cent of male mice treated with endosulfan showed clear signs of infertility. Its impact on mice is an underestimation as we used lower doses. People are exposed to much higher doses and for a longer duration,” said Prof. Sathees Raghavan from the Department of Biochemisty, IISc, Bengaluru and the senior author of the paper.
Ten animals per group were studied and the experiment was repeated three times.
A dose equivalent of 3 mg/kg of body weight was used in the experiment. This dose is equivalent to that seen in places where people live. Compared with 20-700 microgram/litre serum concentration of endosulfan in humans, the treated mice had a serum concentration of just 40-60 microgram/litre.
Besides lower serum concentration, the duration of exposure was also very short. The animals were treated with four doses of endosulfan per day on every alternative day for eight days. In all, each mouse in the treatment group was exposed to 16 doses.
Detailed studies revealed that among all organs, liver, lungs and testes were maximally affected after the very first day of treatment. Kidney, liver and brain were not affected. In the testicles, many seminiferous tubules (where sperms are produced) were completely devoid of mother cells that become sperms.
Since it takes 30-35 days for mother cells to become sperms, at the end of 35 days after the latest treatment, the number of sperms in treated mice was far lower compared with mice that were not treated with the pesticide (control group).
Besides lower sperm count, the motility of the sperms was affected. Twenty-five days after the last day of endosulfan treatment, the motility of sperms was significantly affected. However, at day 35, the pesticide’s effect on motility was not significant. “That’s because spermatogenesis [the process in which spermatozoa are produced from male germ cells] gets over by 30-35 days. So the effect probably wears off after that,” said Prof. Raghavan.
It is to be remembered that unlike mice that were exposed to endosulfan for a limited period, people living in the areas where the pesticide is being used are continually exposed and may see the effect on motility continuing to be significant if effects on humans is the same as seen in mice.
The researchers carried out mating experiments to double check the effect of lower sperm count and less motility in mice. They allowed the pesticide-treated male mice to breed with female mice (in 1:3 ratio of male and females) that were not treated with endosulfan. The mating experiments were carried during three periods (5-10 days, 20-25 days and 30-35 days after treatment with the pesticide). “About 30 per cent of males did not give any progeny,” he said.
“In all three mating windows, the percentage of infertile males increased as opposed to zero infertile males in control groups, indicating that the reduced sperm count and motility contributed towards reduced fertility in males,” notes the paper.
What is indeed significant is that the 20-25-day window showed the most infertile males. This is only to be expected as at 25 days post-treatment, the sperm count had dropped to its lowest point and the sperms had the least motility.
Besides fertility, the pesticide was found to damage the DNA and increase the levels of error prone DNA repair leading to genomic instability. Reactive oxygen species were found to be the culprits causing damaging to DNA and the DNA repair mechanism.
The World Health Organization has classified endosulfan as a Class II pesticide (moderately toxic) and it is listed under the Stockholm convention as a persistent organic pollutant, given its environmental impact. The Stockholm Convention held in 2011 had advised the phasing out of the pesticide over a five-year period and India has agreed to phase out its use by 2017. Currently, Kerala and Karnataka have banned its use as people in Kasargod district in Kerala had reported several health problems related to its use. The pesticide was aerially sprayed on cashew plantations in the district.